RNA interference (RNAi) induced through double-stranded RNA (dsRNA) has been used widely to study gene function in insects. In this paper we demonstrate the efficacy of RNAi in the cotton bollworm, Helicoverpa armigera. Using CYP6B6 as the target gene, which is expressed in the fat baby and midgut of the lepidopteran pest H. armigera, we constructed the vector which expressed dsRNA of CYP6B6. Northern blot analysis showed that dsRNA expressed in the Escherichia coli (HT115) was target gene. The results also showed that the gene expression level and protein expression level of H. armigera larvae fed with dsRNA expressed by E. coli were significantly lower than those of all controls, but the gene expression level was more obvious than that at the protein level; significant lethality differences were also found between HT115 bacteria containing L4440-dsC1 treatment and HT115 bacteria containing L4440 vector or CK (ddH2O) in instar larvae on 4 day when continuous feeding, 32.45% mortality was recorded in the group of feeding HT115 bacteria containing L4440-dsC1 on 10 day. Our results suggest that the RNAi pathway can be exploited to control insect pests.

Many regions are increasingly threatened by agricultural pests but suffer from a lack of data that hampers the development of adequate population dynamics models that could contribute to pest management strategies. Here, we present a new model relating pest survival to temperature and compare its performance with two published models. We were particularly interested in their ability to simulate the deleterious effect of extreme temperatures even when adjusted to datasets that did not include extreme temperature conditions. We adjusted the models to survival data of three species of potato tuber moth (PTM), some major pests in the Tropical Andes. To evaluate model performance, we considered both goodness-of-fit and robustness. The latter consisted in evaluating their ability to predict the actual altitudinal limits of the species in the Ecuadorian Andes. We found that even though our model did not always provide the best fit to data, it predicted extreme temperature mortality and altitudinal limits accurately and better than the other two models. Our study shows that the ability to accurately represent the physiological limits of species is important to provide robust predictions of invasive pests' potential distribution, particularly in places where temperatures approach lethal extremes. The value of our model lies in its ability to simulate accurate thermal tolerance curves even with small datasets, which is useful in places where adequate pest management is urgent but data are scarce.

Among the herbivorous arthropods that feed on strawberry, the most important are the two-spotted spider mite (TSSM), Tetranychus urticae Koch, and several species of aphids. Mites and aphids belong to different guilds that coexist in the field and feed on the undersides of strawberry leaflets. However, the occurrence of large numbers of individuals of both species on the same leaflet is rarely recorded. We hypothesize that negative interactions between TSSM and aphids explain the intraplant distribution of these herbivores. We first examined the spatial coincidence of both herbivores in the field. Under experimental conditions, we then analyzed: (i) the rate of increase of TSSM and the aphid Chaetosiphon fragaefolii (Cockerell), growing individually and together; (ii) the effect of honeydew on TSSM preference; and (iii) the effect of previous strawberry leaflet damage by TSSM on C. fragaefolii preference. The proportion of TSSM that coincided with at least one aphid decreased as the percentage of leaflets with TSSM increased. The spatial coincidence index between aphids and TSSM increased together with the percentage of TSSM-infested leaflets. TSSM showed both a lower rate of increase when they shared the same leaflet with C. fragaefolii and lower fecundity on strawberry discs with honeydew. The rate of increase of C. fragaefolii did not change on co-occupied leaves, but the aphid species moved to the other side of leaflets shared with TSSM. Negative interactions resulting in a tendency for species to avoid each other, such as demonstrated herein, can affect distribution and performance of herbivorous arthropods.

Clones have been in the forefront of biological interest for many years. Even so, open discussions continue to surround the concept of clonality, which has been recently much debated in the scientific literature, both in terms of philosophical meaning as well as empirical determination. Philosophically, the clone is the horizontally produced lineage from a single fertlized egg (e.g. mammals by division of the fertilized egg and representing a single generation) or vertically produced offspring (e.g. aphids representing different successive generations) from a single asexual stem mother (originally for a particular lineage, following hatching of the overwintering sexual egg in the spring); empirically, the aspect of genetic fidelity is also considered important, so-called clones being assumed to have an identical genome among clone mates. In reality of course, such members of a clonal lineage must differ at various regions of the genome, since mutation is a fundamental property of the DNA itself. Yet few studies have so far set out to show this empirically in eukaryotic organisms, which indulge in periods of asexual reproduction, sometimes, as in aphids, over many generations. In the present study, we have investigated asexual lineages of the grain aphid, Sitobion avenae (F.), a global pest of cereals, over five successive generations employing AFLP-PCR molecular techniques. Our main interest was to see how much variation was present in the early generations and if this variation was transmitted through the asexual lineages. By monitoring AFLP-based polymorphisms, we show that, in this aphid species, of a total of 110 individuals from two lineages tested (termed SA and SB), random mutations (band deletions, more rarely additions) were apparent from the third generation onwards, and although some mutations were found to be transmitted transgenerationally, others were rarely transmitted through the particular lineages they were detected in. Using Arlequin v. 2.0, average gene diversity within the lineages was found to be 0.024 ± 0.013 and 0.031 ± 0.016 for SA and SB, respectively. It was also found from the rearing of the lineages that one lineage, SA, was more fecund than the other lineage, SB, over the five generations (N = 818 vs. N = 358 total stem mothers plus nymphs for the two lineages, respectively).

When assessing the benefits and risks of transgenic crops, one consideration is their relative effects on non-target arthropod (NTA) abundance and functions within agroecosystems. Several laboratory and field trials have been conducted in Spain since the late 1990s to assess this issue. A consideration in the design of field trials is whether it is necessary to sample most NTAs living in the crop or only representative taxa that perform main ecological functions and have a good capacity to detect small changes in their abundance. Small changes in the field abundance of an effective representative taxon should be detectable using standard experimental protocols. The ability of a species to reveal differences across treatments may be analysed by examining the detectable treatment effects for surveyed non-target organisms. Analysis of data from several NTAs recorded in 14 field trials conducted over 10 years using complete block designs allowed us to select a number of representative taxa capable of detecting changes in the density or activity of arthropod herbivores, predators, parasitoids and decomposers in transgenic and non-transgenic maize varieties. The most suitable NTA as representative taxa (with detectable treatment effects below 50%) included leafhoppers among arthropod herbivores, Orius spp., Araneae, and Carabidae among predators, chalcidids, particularly the family Mymaridae, among parasitoids and Chloropidae as decomposer. Details of sampling techniques for each sampled taxa and their advantages and disadvantages are discussed. It is concluded that abundance of taxa is the most influential factor determining their capacity to detect changes caused by genetically modified varieties.

Diverse agroecosystems offer phytophagous insects a wide choice of host plants. Myzus persicae is a polyphagous aphid common in moderate climates. During its life cycle it alternates between primary and secondary hosts. A spatial genetic population structure may arise due to environmental factors and reproduction modes. The aim of this work was to determine the spatial and temporal genetic population structure of M. persicae in relation to host plants and climatic conditions. For this, 923 individuals of M. persicae collected from six plant families between 2005 and 2008 in south-eastern Spain were genotyped for eight microsatellite loci. The population structure was inferred by neighbour-joining, analysis of molecular variance (AMOVA) and Bayesian analyses. Moderate polymorphism was observed for the eight loci in almost all the samples. No differences in the number of alleles were observed between primary and secondary hosts or between geographical areas. The proportion of unique genotypes found in the primary host was similar in the north (0.961 ± 0.036) and the south (0.987 ± 0.013), while in the secondary host it was higher in the north (0.801 ± 0.159) than in the south (0.318 ± 0.063). Heterozygosity excess and linkage disequilibrium suggest a high representation of obligate parthenogens in areas with warmer climate and in the secondary hosts. The FST-values pointed to no genetic differentiation of M. persicae on the different plant families. FST-values, AMOVA and Bayesian model-based cluster analyses pointed to a significant population structure that was related to primary and secondary hosts. Differences between primary and secondary hosts could be due to the overrepresentation of parthenogens on herbaceous plants.

β-Aminobutyric acid (BABA) is known to induce resistance to microbial pathogens, nematodes and insects in several host plant/pest systems. The present study was undertaken to determine whether a similar effect of BABA occurred against the Asian citrus psyllid, Diaphorina citri Kuwayama, in citrus. A 25 mM drench application of BABA significantly reduced the number of eggs/plant as compared with a water control, whereas 200 and 100 mM applications of BABA reduced the numbers of nymphs/plant and adults/plants, respectively. A 5 mM foliar application of BABA significantly reduced the number of adults but not eggs or nymphs when compared with a water control treatment. In addition, leaf-dip bioassays using various concentrations (25–500 mM) of BABA indicated no direct toxic effect on 2nd and 5th instar nymphs or adult D. citri. BABA-treated plants were characterized by significantly lower levels of iron, magnesium, phosphorus, sodium, sulfur and zinc as compared with control plants. The expression level of the PR-2 gene (β-1,3-glucanase) in BABA-treated plants that were also damaged by D. citri adult feeding was significantly higher than in plants exposed to BABA, D. citri feeding alone or control plants. Our results indicate the potential for using BABA as a systemic acquired resistance management tool for D. citri.

The cryptic species Middle East-Asia Minor 1 (MEAM1), formerly referred to as ‘B biotype’, of the whitefly Bemisia tabaci complex entered China in the mid 1990s, and the Mediterranean (MED) cryptic species, formerly referred to as ‘Q biotype’, of the same whitefly complex entered China around 2003. Field surveys in China after 2003 indicate that in many regions MED has been replacing the earlier invader MEAM1. The factors underlying this displacement are unclear. We conducted laboratory experiments and field sampling to examine the effects of insecticide application on the competitive interactions between MEAM1 and MED. In the laboratory, on cotton, a plant showing similar levels of suitability to both whitefly species, MEAM1 displaced MED in five generations when initial populations of the two species were equal and no insecticide was applied. In contrast, MED displaced MEAM1 in seven and two generations, respectively, when 12.5 and 50.0 mg l−1 imidacloprid was applied to the plants via soil drench. Field sampling indicated that in a single season MED displaced MEAM1 on crops heavily sprayed with neonicotinoid insecticides but the relative abundance of the two species changed little on crops without insecticide spray. We also examined the effects of host plants on the competitive interactions between the two species in the laboratory. When cohorts with equal abundance of MEAM1 and MED were set up on different host plants, MEAM1 displaced MED on cabbage and tomato in five and seven generations, respectively, but MED displaced MEAM1 on pepper in two generations. As field populations of MED have lower susceptibility than those of MEAM1 to nearly all commonly used insecticides including imidacloprid, insecticide application seems to have played a major role in shifting the species competitive interaction effects in favour of MED in the field across China. Host plants may also shape competition between the two species depending on the relative levels of plant suitability.

The Holarctic genus Reticulitermes shows seven species within the Mediterranean Basin. While phylogeny and systematics at continental level has been deeply investigated, a few studies concentrated on local ranges. To gain a clearer picture of the diversity and evolution of the Italian species Reticulitermes lucifugus, we analyzed the mitochondrial cytochrome oxidase II (COII) gene marker in newly collected colonies across the Peninsula. Data were gathered with all R. lucifugus sequences available from previous studies; COII sequences of the closely related Iberian taxa were also added to the data set. Maximum-likelihood, median-joining and statistical parsimony network elaborations on the resulting 119 colonies all agreed in indicating that: (i) the Sardo-Corsican subspecies R. lucifugus corsicus, strictly related to Southern Italian populations (including the Sicilian ones), is phylogenetically closer to the Iberian Reticulitermes grassei; and (ii) R. lucifugus lucifugus peninsular populations are structured into three clusters. The phylogenetic relationships and the biogeography of extant taxa suggest a scenario in which R. lucifugus ancestors colonized the Italian region through the Sardo-Corsican microplate during its Oligocene-Miocene anticlockwise rotation. Moreover, well after the colonization took place, northward range expansion might have produced the presently observed genetic diversity, as inferred from haplotype and nucleotide diversity estimates. On the whole, this study highlights the evolution of Italian Reticulitermes taxa and supports the importance of a wide taxon sampling especially when dealing with organisms easily dispersed by human activities.

Rhynchophorus ferrugineus (Coleoptera, Curculionidae) is the most threatening pest of palms worldwide. The potential of gamma-irradiated males to spread a pathogenic strain of the entomopathogenic fungus Beauveria bassiana (Ascomycota: Clavicipitaceae) to control this pest was studied. First, the effects of gamma irradiation (15 and 25 Gy) on the mating success and performance of adult males irradiated at age one day were studied in the laboratory. Although male longevity decreased after irradiation (118.6 vs. 244.7 days for irradiated and control males, respectively) and their testes suffered from the treatment, fecundity of mated females did not depend on the irradiation status of the male (86.8 ± 5.5 eggs in 15 days). However, egg hatching was significantly lower in couples with irradiated males (31.4% vs. 86.5% for irradiated and control couples, respectively), and this value decreased after a second mating (6.1% vs. 85.9%). Therefore, irradiation did not affect male sexual competiveness but sperm quality. Second, a semi-field assay was carried out to evaluate infestation in young Phoenix canariensis caused by different combinations of couples with irradiated and/or B. bassiana-challenged males. The number of immature stages found in infested palms was significantly higher when females mated with untreated males and lower when mated with irradiated males (either B. bassiana-infected or not). Some females from the fungus-challenged treatments showed post-mortem hyphal growth, and this horizontal transmission proves that irradiated males could act as a vector for B. bassiana and should be considered as a new method to improve the biological control of R. ferrugineus.

The ladybird beetle, Eriopis connexa (Germar) (Coleoptera: Coccinellidae), is one of the commonest predators of aphids (Hemiptera: Aphididae) in the cotton agroecosystem and in many other row and fruit crops in Brazil, and has been introduced into other countries such as the USA for purposes of aphid control. In addition, the boll weevil, Anthonomus grandis Boheman (Coleoptera: Curculionidae) is the most serious cotton pest where it occurs, including Brazil. Controlling boll weevils and other pests such as cotton defoliators still tends to involve the intense application of insecticides to secure cotton production. The pyrethroid insecticide lambda-cyhalothrin (LCT) is commonly used, but this compound is not effective against aphids; hence, a desirable strategy would be to maintain E. connexa populations in cotton fields where LCT is applied. Using populations of E. connexa resistant (Res) and susceptible (Sus) to LCT, we compared behavioural responses on treated cotton plants and under confinement on partially and fully treated surfaces, and assessed the insects' survival on treated plants compared with that of the boll weevil. The E. connexa resistant population caged on treated plants with 15 and 75 g a.i. ha−1 exhibited ≫82% survival for both insecticide concentrations compared with ≪3% and ≪17% survival for susceptible E. connexa populations and boll weevils, respectively. The response of E. connexa Res and Sus populations when released, either on the soil or on the plant canopy, indicated avoidance towards treated plants, as measured by elapsed time to assess the plant. When compared with susceptible individuals, resistant ones took longer time to suffer insecticide knockdown, had a higher recovery rate after suffering knockdown, and spent more time in the plant canopy. Based on behavioural parameters evaluated in treated arenas, no ladybird beetles exhibited repellency. However, irritability was evident, with the susceptible population exhibiting greater irritability compared with the resistant population and a subgroup comprising resistant individuals that had recovered from knockdown. The outcomes for the E. connexa Res population indicate a promising strategy for its maintenance when using the insecticide LCT in integrated pest management schemes to control boll weevil or other non-target pest of ladybird beetles in cotton fields.

Two new approaches were examined, aimed at increasing survival of the house fly (Musca domestica L.) larvae under mass-rearing conditions of a biodegradation facility: modification of the larval substrate and dispersal of the eggs during inoculation. The two types of pig manure used in this study (manure with sawdust and manure without sawdust) differed in terms of larval survival and nutritional value for the house fly larvae. Larval survival in manure without sawdust in the control treatment was low (46.8 ± 2.1%) and its nutritional value for the larvae were high. Addition of 5.7% of previously biodegraded manure did not significantly affect larval survival (52.3 ± 1.9%), but larval development was faster and the pupae were significantly smaller (14.28 ± 0.4 mg) compared to the control (16.29 ± 0.5 mg). Using alternative substrate for incubation of eggs and first-instar larvae significantly increased larval survival (63.3 ± 3.3%) and decreased the mean weight of produced pupae (14.39 ± 0.71 mg). Overall, the weight of recovered biomass in the alternative substrate treatment increased by 14.3 kg ton−1 of manure compared to the control. Larval survival in manure with sawdust was generally higher than 70%, but its nutritional value for the larvae was lower than in manure without sawdust. Dispersal of eggs over the surface of manure with sawdust significantly affected larval survival and mean weight of pupae. Larval survival was significantly lower (59.2 ± 4.0%) and pupae were significantly heavier (18.45 ± 0.8 mg) when eggs were applied to a small area on the manure surface (spot treatment), as compared to diagonal, Z-line and multiple zig-zag dispersal (72.5 ± 2.4 to 74.6 ± 3.0% and 14.76 ± 0.6 to 15.97 ± 0.6 mg, respectively). No significant differences were observed in larval survival or mean weight of pupae when comparing the diagonal, Z-line and multiple zig-zag dispersal patterns. Implementation of the techniques which improve larval survival and increase the weight of produced fly biomass may decrease demand for production of house fly eggs and, therefore, reduce the maintenance costs of adult colony, as well as increase the revenue earned by selling the products.

Rapid, precise and timely identification of invasive pest insects such as aphids is important and a challenge worldwide due to their complex life cycles, parthenogenetic reproduction, sex and colour morphs. In this respect, DNA barcoding employing a 658 bp fragment of 5′ region of the mitochondrial cytochrome oxidase I (CO-I) gene is an effective tool in addressing the above. In the present study, we employed CO-I for discriminating 142 individuals representing 32 species of aphids from India. Sequence analyses revealed that the intraspecific and interspecific distances ranged from zero to 3.8% and 2.31 to 18.9%, respectively. In addition, the study also showed for the first time the prevalence of three cryptic species, namely Brevicoryne brassicae (Linnaeus), Hyperomyzus carduellinus (Theobald) and Brachycaudus helichrysi (Kaltenbach) from India. Our work has clearly demonstrated that DNA barcoding is an efficient and accurate method for identification of aphid species (including cryptic species), an approach that potentially could play an important role in formulating viable pest management strategies, more especially biocontrol.

Epitrix species (Coleoptera: Chrysomelidae) feed mostly on plants from the family Solanaceae and some of them are major pests of potato crops. All Epitrix species are morphologically highly similar, which makes them difficult to identify and limits their study and management. Identification of species is mostly based on the observation of the genitalia and requires a high level of expertise. Here, we propose a tool to reliably identify all developmental stages of the most economically important Epitrix species feeding on potato in Europe and North America (Epitrix cucumeris, Epitrix similaris, Epitrix tuberis, Epitrix subcrinita and Epitrix hirtipennis). We first sequenced two DNA markers (mitochondrial cytochrome c oxidase I (COI) and nuclear internal transcribed spacer 2 (ITS2)) to test their effectiveness in differentiating among six Epitrix species (126 specimens). Morphospecies of Epitrix were well-differentiated by both DNA barcodes and no mitochondrial introgression was detected. Then, we developed an RFLP-based diagnostic method and showed that unambiguous species discrimination can be achieved by using the sole restriction enzyme TaqI on COI polymerase chain reaction products. The tool proposed here should improve our knowledge about Epitrix species biology, distribution and host range, three capacities that are particularly important in the detection and management of these pest species. Specifically, this tool should help prevent the introduction of E. tuberis and E. subcrinita in Europe and limit the spread of the recently introduced E. cucumeris and E. similaris, with minimal disruption to Solanaceae trade.

Following the global uptake of transgenic cotton several Hemipteran pests have emerged as primary targets for pesticide control. Previous research on one such emergent pest: the green mirid, Creontiades dilutus, indicated differential use of two crop hosts, cotton (Gossypium hirsutum, Malvaceae) and lucerne (alfalfa) (Medicago sativa, Fabaceae). We tested the hypothesis that this apparent demographic independence of lucerne and cotton inhabiting mirids is the result of cryptic species being associated with these two crops. We assessed gene flow using microsatellite markers across adjacent cotton and lucerne crops at three geographically separated sites (up to 900 km apart). We also analysed the recent feeding behaviour of these insects by amplifying chloroplast markers from their gut contents. We find high gene flow between these two crops (mean pair wise FST between host plants=0.0141 within localities), and no evidence of cryptic species. Furthermore, the gut analyses revealed evidence of substantial recent movement between these two crops. We discuss the implications of these results for interpreting multiple host use in this species and setting future research priorities for this economically important pest.